The balance between activated Th1 and Th2 lymphocytes plays a central role in the regulation of the allergic responses, and has particularly marked effects on IgG subclasses and IgE production. We have recently shown that i.d gene vaccination induces a polarized Th1 response which inhibits subsequent induction of Th2 response and could down-regulate an on-going Th2 response in an antigen specific fashion. This was shown by a substantial fall in total and specific IgE levels, an increase in IgG2a titers, an increase in IFNgamma and a decrease in IL-5 production by antigen specific CD4+ Th cells as well as by inhibiting the late phase reaction (LPR) of the allergic response. In subsequent experiments we found that the presence of immunostimulatory DNA sequence (ISS) in the plasmid (p) backbone is required for the observed Th1 response. The ISS activate the innate immune system to release IFNalpha, IFNbeta, IL-12 and IL-18, cytokines which promote the differentation of Th1 lymphocytes. Furthermore, we could exploit this information to induce a Th1-like response and to inhibit IgE synthesis by co-administration of a protein antigen with non-coding, ISs enriched pDNA. The objective of this proposal is to establish the principles of allergen gene vaccination in vio. We will immunize mice with different types of pDNA constructs and screen their ability to induce a Th1 response (e.g. via their ISS) to the gene product (allergen) and their differential effectiveness in diverting an on-going Th2 (allergic) response to a Th1 response. This immune modulation will be assessed by measurements of cytokine release, IgG isotypes formation and by IgE antibody inhibition. The ability of gene vaccination to inhibit or down-regulate the LPR will be also determined in a lung model of allergic inflammation. The immunological, histological and pulmonary responses to the inhaled allergen will then be assessed. The results obtained from the proposed experiments will increase our knowledge and insight in manipulating the immune response by gene vaccination and will eventually lead to phase 1 clinical studies in allergic patients. Thus, by affecting the immediate (IgE) and the late phase reactions, allergen gene vaccination potentially offer a new approach for the therapy of allergic diseases in humans.